[0001] This invention relates to a process for producing gelatin spherical gels, and more
particularly to a process for producing polypeptide spherical gels suitable for an
embolic agent or a carrier for impregnating pharmaceuticals.
[0002] In recent years, studies on embolization of arteriae, selective intra-arterial injection
of drugs, and the like have been made extensively in the field of radiology. These
studies have their objects to conduct embolization on a trophic arteria having neoplasm
as pretreatment of surgical removal of the neoplasm so as to reduce the bleeding amount
during operation, or to reduce the neoplasm by closure of tropic arteria or by selective
injection of pharmaceuticals from tropic arteria in the case of carcinomatous neoplasm
when radical operation is impossible.
[0003] On the other hand, embolic agents can be divided into two groups depending on purposes,
that is, those having ephemeral embolism or having solubility and those having permanent
embolism or having insolubility. As embolic agents for such purposes, there are used
those obtained by dissolving gelatin in water, foaming, subjecting to lyophilization
to.form spongy body and cutting into several millimeters at a side or powdered; those
obtained by adding formaldehyde to gelatin, conducting polymerization with heating,
blowing air bubbles thereinto to form spongy body and cutting into several millimeters
at a side or powdered; or the like. In the case of selective injection of pharmaceuticals,
these substances are impreganted with pharmaceuticals before use. But since these
substances are in broken form, they have a defect in that they are difficult to adhere
to blood vessels compared with spherical ones.
[0004] As spherical substances available commercially, there are known those made from organic
polymers such as polystyrenes, poly(acrylic acid ester)s, polyvinyl alcohols, those
made from inorganic substances such as silica, glass, etc. But these substances have
a problem from the viewpoint of safety when they are included in a living body.
[0005] It is an object of this invention to provide a process for producing spherical substances
usable as embolic agent, etc., overcoming the problem mentioned above.
[0006] This invention provides a process for producing gelatin spherical gels which comprises
dispersing an aqueous solution of gelatin in a dispersing medium obtained by dissolving
water-insoluble ethyl cellulose in a non-polar organic solvent which is not miscible
with water, and removing the water by distillation.
[0007] Gelatin has various molecular weights depending on raw materials used. In this invention,
there is no particular limit to the molecular weight.
[0008] The concentration of gelatin in an aqueous solution of gelatin is preferably 5% by
weight or more and the saturated concentration or lower. When the gelatin concentration
is too low, the productivity is lowered.
[0009] In a dispersing medium, an aqueous solution of gelatin and one or more crosslinking
agents, if desired, may be dispersed and subjected to a crosslinking reaction. This
can be conducted by adding the aqueous solution of gelatin and crosslinking agent
dissolved in water to a liquid which is immiscible with water and stirring the resulting
mixture. But since the gelatin easily reacts with the crosslinking agent, it is preferable
to disperse an aqueous solution of gelatin in the liquid immiscible with water, followed
by addition of the crosslinking agent or an aqueous solution thereof thereto.
[0010] Gelatin has functional groups such as -NH
2, -OH and -COOH which become reactive sites at the crosslinking reaction.
[0011] As the crosslinking agent, there can be used a water-soluble compound which can conduct
a crosslinking reaction with gelatin. Examples of the crosslinking agent having reactivity
with the functional groups (e.g. -NH
2 group, -COOH.group) of gelatin are aliphatic dials such as glyoxal, propanedial,
butanedial, pentanedial (glutaraldehyde), hexanedial, etc.; water-soluble polyvalent
epoxides such as ethylene glycol diglycidyl ether, polyethylene glycol diglycidyl
ether, glycerol polyglycidyl ether, diglycerol polyglycidyl ether, sorbitol polyglycidyl
ether, diglycidyl methylhydantion, etc.
[0012] In general, when the amount of crosslinking agent is less than 5/100 equivalent weight
per equivalent weight of reactive functional groups of gelatin (such as -NH
2, -COOH), the crosslinking becomes insufficient to maintain the gelatin particles
in spherical form. Even in such a case, when the water is removed by distillation
according to this invention, the resulting gelatin particles can remain in the spherical
form. If the water is not removed by distillation, the resulting particles are easily
paste-likely agglomerated after the separation and washing of the particles with water.
As the crosslinking agent, it is preferable to use those which do not react with ethyl
cellulose.
[0013] The dispersing medium usable in this invention is a compound which is not miscible
with water, can dissolve the water-insoluble ethyl cellulose, and can form an azeotropic
mixture with water or has a higher boiling point than water. Such a compound is a
non-polar organic solvent such as an alicyclic hydrocarbon, and the like. The non-polar
organic solvent can be used together with one or more polar solvents such as esters,
ketones, halogenated alkanes, ethers, alcohols or aromatic hydrocarbons, these being
unreactive with the crosslinking agent.
[0014] Preferable examples of the alicyclic hydrocarbons are those having 5 to 10 carbon
atoms or alkyl substituted alcyclic hydrocarbons such as cyclopentane, cyclohexane,
cycloheptane, methylcyclohexane, cyclooctane, decalin, etc. These alicyclic hydrocarbons
can be used alone or as a mixture thereof.
[0015] The aromatic hydrocarbons include aromatic hydrocarbons and halogen substituted aromatic
hydrocarbons having 6 to 8 carbon atoms. Examples of these aromatic hydrocarbons are
benzene, toluene, xylenes, ethylbenzene, chlorobenzene, dichlorobenzene, bromobenzene,
dibromobenzene, etc. These aromatic hydrocarbons can be used alone or as a mixture
thereof.
[0016] As the esters, there can preferably be used those obtained from fatty acids with
1 to 8 carbon atoms or aromatic carboxylic acids with 7 to 8 carbon atoms and alcanols
having 1 to 8 carbons, alone or as a mixture thereof. Examples of the esters are methyl
acetate, ethyl acetate, n-butyl acetate, benzyl acetate, methoxyethyl acetate, methyl
caproate, methyl benzoate, diethyl phthalate, etc.
[0017] As the ketones, there can preferably be used aliphatic ketones having 5 to 8 carbon
atoms, and aromatic ketones having 8 to 13 carbon atoms, alone or as a mixture thereof.
Examples of the ketones are methyl isobutyl ketone, cyclohexanone, methyl amyl ketone,
hexyl methyl ketone, acetophenone, benzophenone, etc.
[0018] As the halogenated alkanes, there can preferably be used alkanes having 1 to 4 substituted
halogens alone or as a mixture thereof. Examples of the halogenated alkanes are methylene
chloride, carbon tetrachloride, 1,2-dichloroethane, 1,1,2-trichloroethane, pentachloroethane,
1,2-dichloropropane, 1,2-dichlorbutane, 1,2-dibromoethane, etc.
[0019] As the ethers, there can preferably be used straight-chain or cyclic ethers having
4 to 8 carbon atoms alone or as a mixture thereof. Examples of the ethers are di-n-propyl
ether, di-n-butyl ether, tetrahydrofuran, dioxane, diethylene glycol dimethyl ether,
etc.
[0020] As the alcohols, there can preferably be used alkanols having 4 to 8 carbon atoms
or alkoxy alkanols having 4 to 8 carbon atoms alone or as a mixture thereof. Examples
of the alcohols are n-butanol, n-octanol, methoxyethanol, ethoxyethanol, butoxyethanol,
diethylene glycol monomethyl ether, etc.
[0021] As the water-insoluble ethyl cellulose, there can be used those having an ethoxy
group content of 43 to 50%. The words "ethoxy group content" mean a weight percent
of the ethoxy group in ethyl cellulose. Among those mentioned above, those having
relatively smaller ethoxy group content, for example, those having 43 to 46% by weight
of the ethoxy group content, are hardly soluble in the above-mentioned non-polar organic
solvent. Therefore, when the above-mentioned non-polar organic solvent is used alone
as the dispersing medium, it is preferable to use those having 47 to 50% by weight
of the ethoxy group content.
[0022] When the water-insoluble ethyl cellulose is used in combination with one or more
above-mentioned non-polar organic solvents such as alicyclic hydrocarbons, there can
be formed a particularly preferable dispersing medium. But since the solubility of
ethyl cellulose in a non-polar organic solvent is poor as mentioned above, it is preferable
to use the above-mentioned polar solvent or aromatic hydrocarbon together as dispersing
medium.
[0023] When a mixture of non-polar organic solvent and at least one aromatic hydrocarbon
is used as dispersing medium, the aromatic hydrocarbon can be used in an amount of
preferably 400% by weight or less, more preferably 200% by weight or less based on
the weight of the non-polar organic solvent.
[0024] When a mixture of non-polar organic solvent and at least one of the esters, ketones
and halogenated alkanes is used as dispersing medium, the latter component can be
used in an amount of preferably 200% by weight or less, more preferably 150% by weight
or less, based on the weight of the non-polar organic solvent.
[0025] When a mixture of non-polar organic solvent and at least one of the ethers and alcohols
is used as dispersing medium, the latter component can be used in an amount of preferably
30% by weight or less, more preferably 20% by weight or less, based on the weight
of the non-polar organic solvent.
[0026] When the aromatic hydrocarbons, ketones, esters, halogenated alkanes, ethers, alcohols
are used together with the non-polar organic solvent in amounts larger than the values
mentioned above, there is a tendency not to provide a good dispersing medium.
[0027] The water-insoluble ethyl cellulose can be used in an amount of preferably 0.05 to
10% by weight, more preferably 0.5 to 5% by weight, based on the total weight of the
dispersing medium.
[0028] The dispersing medium used in this invention obtained by dissolving the water-insoluble
ethyl cellulose in the water-immiscible organic dispersing medium can disperse non-crosslinked
gelatin effectively.
[0029] It is preferable to use the dispersing medium in an amount of preferably 50 to 2000%
by weight, more preferably 100 to 1000% by weight, based on the total weight of gelatin,
the crosslinking agent and water (hereinafter referred to as "the amount of water
phase"). When the amount of the dispersing medium is too much, the productivity becomes
poor, while when the amount of the dispersing medium is too small, the stability of
dispersing medium becomes poor.
[0030] In order to disperse the aqueous solution of gelatin in the dispersing medium, there
can be employed a method of mixing the two with stirring, a method of adding the aqueous
solution of gelatin to the dispersing medium with stirring, and the like. As the stirring
method, there can be employed a stirring method by using an emulsator accompanying
high-speed shearing, a stirring method by using a marine propeller stirrer or magnetic
stirrer not accompanying mechanical cutting or grinding, and the like. These stirring
methods can be selected depending on the desired particle size.
[0031] It is preferable to conduct the dispersion of the aqueous solution of gelatin at
a temperature from room temperature to the boiling point of water or the dispersing
medium. After the completion of dispersion, the water is removed by distillation at
elevated temperatures or under reduced pressure.
[0032] After removal of the water by distillation, the resulting gelatin spherical particles
can be recovered by filtration or decantation. The solvent which forms an azeotropic
mixture with water can be removed by distillation together with the distillation of
water. In such a case, when the amount of dispersing medium becomes too small, such
a solvent can be supplemented during the distillation. When the washing of the resulting
particles is conducted by using a solvent having a relatively low boiling point and
the drying is conducted under atmospheric or reduced pressure, the particles can be
purified while maintaining the spherical form.
[0033] If a very limited range of particle size is required, a conventional classification
can be employed.
[0034] This invention is illustrated by way of the following Examples and Comparative Example.
Example 1
[0035] In a dispersing medium comprising 150 g of decalin and 50g of toluene, 6 g of ethyl
cellulose (the ethoxy group content: 49% by weight) was dissolved. The resulting dispersing
medium was introduced into a 500-ml flask equipped with a cooling tube and agitating
blades made from polytetrafluoroethylene. The stirring speed was controlled at 400
r.p.m. and the temperature was raised to 70°C. Then, 40 g of a 30% by weight aqueous
solution of gelatin dissolved at 50°C was added to the flask, wherein the stirring
was continued for 5 minutes to produce spherical particles. Then, the temperature
was raised to 110°C to remove the water by distillation. The resulting particles were
collected by filtration, washed with ethyl acetate, followed by washing with acetone.
The particles had a spherical form and a particle size of 0.1 to 1 mm when observed
by a microscope.
Example 2
[0036] In a dispersing medium of 200 g of decalin, 6 g of ethyl cellulose (the ethoxy group
content: 49% by weight) was dissolved. The resulting dispersing medium was introduced
into a 500-ml flask equipped with a cooling tube and agitating blades made from polytetrafluoroethylene.
The stirring speed was controlled at 400 r.p.m. and the temperature was raised to
70°C. Then, 40 g of a 30% by weight aqueous solution of gelatin dissolved at 50°C
was added to the flask, wherein the stirring was continued for 5 minutes to produce
spherical particles. Then, the temperature was raised to 110°C to remove the water
by distillation. The resulting particles were collected by filtration, washed with
ethyl acetate, followed by washing with acetone. The particles had a spherical form
and had a particle size of 0.1 to 1 mm when observed by a microscope.
Example 3
[0037] In a dispersing medium comprising 150 g of decalin and 50 g of toluene, 6 g of ethyl
cellulose (the ethoxy group content: 49% by weight) was dissolved. The resulting dispersing
medium was introduced into a 500-ml flask equipped with a cooling tube and agitating
blades made from polytetrafluoroethylene. The stirring speed was controlled at 400
r.p.m. and the temperature was raised to 70°C. Then, 40 g of a 30% by weight aqueous
solution of gelatin dissolved at 50°C was added to the flask, and 0.9 g of a 5% by
weight aqueous solution of glutaraldehyde (9/200 equivalent weight of glutaraldehyde
per equivalent weight of the amino group of gelatin) was also added to the flask,
wherein the stirring was continued for 5 minutes to produce spherical particles. Then,
the temperature was raised to 110°C to remove the water by distillation. The resulting
particles were collected by filtration, washed with ethyl acetate, followed by washing
with acetone. The particles had a spherical form and had a particle size of 0.1 to
1 mm when observed by a microscope.
Comparative Example 1
[0038] In a dispersing medium comprising 150 g of decalin and 50 g of toluene, 6 g of ethyl
cellulose (the ethoxy group content: 49% by weight) was dissolved. The resulting dispersing
medium was introduced into a 500-ml flask equipped with a cooling tube and agitating
blades made from polytetrafluoroethylene. The stirring speed was controlled at 400
r.p.m. and the temperature was raised to 70°C. Then, 40 g of a 30% by weight aqueous
solution of gelatin dissolved at 50°C was added to the flask, wherein the stirring
was continued for 5 minutes to produce spherical particles. Then, the particles were
collected by filtration without removal of the water by distillation, washed with
ethyl acetate, followed by washing with acetone. The resulting particles were agglomerated
to form a paste-like solid.
[0039] As mentioned above, this invention can provide safe gelatin spherical particles.
1. A process for producing gelatin spherical gels which comprises dispersing an aqueous
solution of gelatin in a dispersing medium obtained by dissolving water-insoluble
ethyl cellulose in a non-polar organic solvent which is not miscible with water, and
removing the water by distillation.
2. A process according to Claim 1, wherein the non-polar organic solvent which is
not miscible with water is an alicyclic hydrocarbon.
3. A process according to Claim 2, wherein the alicyclic hydrocarbon is at least one
member selected from the group consisting of cyclopentane, cyclohexane, cycloheptane,
methylcyclohexane, cyclooctane and decalin.
4. A process according to Claim 2 or claim 3, wherein the alicyclic hydrocarbon is
used together with at least one polar solvent or an aromatic hydrocarbon.
5. A process according to Claim 4, wherein the polar solvent is an ester, a ketone,
a halogenated alkane, an ether or an alcohol.
6. A process according to Claim 4, wherein the aromatic hydrocarbon is benzene, toluene,
xylene, ethylbenzene, chlorobenzene, dichlorobenzene, bromobenzene or dibromobenzene.
7. A process according to any preceding Claim /1 wherein the water-insoluble ethyl cellulose has the ethoxy group content of 43 to
50% by weight.
8. A process according to any preceding Claim, wherein the aqueous solution of gelatin
has a concentration of 5% by weight or more.
9. A process according to any preceding Claim wherein the aqueous solution of gelatin
further contains a crosslinking agent.
10. A process according to Claim 9, wherein the crosslinking agent is an aliphatic
dial or a water-soluble polyvalent epoxide.